Anticonvulsant compositions containing diphenylhydantoin compounds and method of using same

ABSTRACT

Various derivatives of barbituric acid and of diphenylhydantoin are described in which alkoxymethyl or benzyloxymethyl groups are attached to the nitrogen atoms of the rings. Therapeutic compositions containing these compounds and their use as anticonvulsant agents are described.

United States Patent Samour Dec. 2, 1975 ANTICONVULSANT COMPOSITIONS 581 Field of Search 424/273 CONTAINING DIPHENYLHYDANTOIN COMPOUNDS AND METHOD OF USING [56] References Cited SAME Inventor: Carlos M. Samour, Wellesley, Mass.

Assignee: The Kendall Company, Walpole,

Mass.

Filed: May 24, 1974 Appl. No.: 472,983

Related U.S. Application Data Division of Ser. No. 125,813, March 18, 1971, which is a division of Ser. NO. 888,943, Dec. 29, 1969, which is a continuation-in-part of Ser. No. 749,972, Aug. 5, 1968, abandoned.

U.S. C1. 424/273; 424/254 Int. C1. A61K 31/415 OTHER PUBLICATIONS Wilson & Grisvold, Textbook of Organic, Medicinal and Pharmaceutical Chemistry, 4th Ed., 1962, (pp. 364-366).

Primary ExaminerLeonard Schenkman [5 7] ABSTRACT 10 Claims, N0 Drawings ANTICONVULSANT COMPOSITIONS CONTAINING DIPI-IENYLI-IYDANTOIN COMPOUNDS AND METHOD OF USING SAME This application is a division of application Ser. No. 125,813, filed Mar. 18, 1971, which is in turn a division of application Ser. No. 888,943, filed Dec. 29, 1969, which in turn is a continuation in part of application Ser. No. 749,972, filed Aug. 5, 1968, now abandoned,

This invention relates to selected substituted barbituric acids and hydantoins, to the use thereof as anticonvulsant agents for treating convulsions and seizures in warm-blooded animals, and to therapeutic compositions containing these compounds. More particularly, this invention relates to l,3-dialkoxy(or dibenzyloxy)- methyl-5,5-disubstituted barbituric acids having the formula wherein R and R arephenyl and ethyl respectively, or R and R are individually alkyl or alkenyl each having 2 to 5 carbon atoms or cycloalkenyl having 5 to 7 carbon atoms, and wherein R and R are individually alkyl having 1 to 12 carbon atoms or benzyl; and to 3-alkoxymethyl-S,S-diphenylhydantoin or 1,3-dialkoxymethyl- 5,5-diphenylhydantoin or 3-benzyloxymethyl-5,5- diphenylhydantoin.

Phenobarbital and diphenylhydantoin have both long been known to be useful as anticonvulsant agents in warm-blooded animals, having been employed in the treatment of epilepsy; however, phenobarbital suffers from the disadvantage that it exhibits hypnotic activity as well as anticonvulsant activity, while diphenylhydantoin has the disadvantage that although not hypnotic, it does have a multiplicity of undesirable side effects, for example, hypertrophic gingivitis, megaloblastic anemia, toxic psychoses, and hirsutism. It has now been found that N-lower alkoxymethyl phenobarbital and N-lower alkoxymethyl diphenylhydantoin compounds are effective anticonvulsants which possess unexpected advantages from the pharmacological standpoint over their respective parent compounds.

Although it has been reported in Doran, Medicinal Chemistry, New York 1959, Vol. IV, John Wiley & Sons, Inc. page 187 (War Department Army Medical Library, Microfilm No. 1720, LG. Farbenindustrie Plant, Elberfeld, Germany) that a material identified by the structural formula of N,N'- dimethoxymethyl phenobarbital has been tested as a hypnotic and found to be without effect, there'has been no indication that it has any useful therapeutic effect and no suggestion as to how to make it or what its properties are.

The compounds of the present invention are effective anticonvulsant agents for convulsions and seizures in warm-blooded animals andare free from hypnotic activity as hereinafter. described. The compounds in which the alkoxygroups are methoxy or ethoxy are preferred; N,N-dimethoxymethyl phenobarbital, and

2 3-methoxymethyl diphenylhydantoin are particularly preferred.

The compounds may be made by reacting an appropriate (e.g. alkali metal salt) barbiturate, such as phenobarbital sodium, or diphenylhydantoin sodium, with chloromethyl or other halomethyl alkyl or benzyl ether in a suitable diluent such as dimethyl formamide, dimethyl sulfoxide, dimethyl acetamide, hexamethylphosphoramide, and the like over a wide range of temperatures, conveniently from about -10C. up to the boiling point of the diluent. The desired products are obtained in high yield and in an excellent state of purity; they can conveniently be isolated by conventional techniques such as extraction, distillation, filtration, preparative chromatography, etc.

Illustrative substituted barbituric acids which are employed in salt form to prepare compounds of the present invention include the following commercially available compounds: barbital, phenobarbital, probarbital,

aprobarbital, amobarbital, allobarbital, vinbarbital, pentobarbital, cyclobarbital, S-allyl- 5-(2-cyclopentenyl )barbituric acid, S-ethyl-S- cycloheptenyl barbituric acid, etc. Diphenylhyydantoin sodium may also be used.

Among the halomethyl alkyl or benzyl ethers suitable for use in the preparation of the compounds of the present invention are chloromethyl methyl ether, chloromethyl ethyl ether, chloromethyl butyl ether, chloromethyl n-dodecyl ether, chloromethyl benzyl ether, and the like.

The compounds of the invention can be formulated with conventional physiologically acceptable vehicles and carriers to make syrups, isotonic solutions, tablets and other dosage forms. Toxicity and effectiveness of the compounds are such that each dosage unit can contain from 5 to 500 mg of active material. However, in view of the high anticonvulsant activity of these compounds, dosage units containing from 5 to 200 mg of the compound are preferred.

The procedures employed for demonstrating efficacy of the compounds as set forth in the following examples were as follows.

All tests, with the exceptions noted, were conducted on adult albino male mice (Charles River strain); the dosage consisted of the active agent suspended in 10% aqueous acacia and was administered orally unless otherwise indicated.

Acute oral toxicity and acute intraperitoneal toxicity were determined in the conventional manner. The results were expressed as L0 the dose required to produce death in 50% of the animals treated, determined graphically, with the limits shown in parentheses.

The dosage required to produce a neurological deficit was determined by the method of Swinyard et al., J. Pharmacol. Exptl. Therap. 106, 319 1952) except that one additional test was employed: the ability of the animal to remain for at least 1 minute on a rotorod, a horizontal rod rotated at 6 rpm. The results were expressed as TD the dose required to produce the effect, determined graphically, with 95% limits.

Anticonvulsant effectiveness of each agent against maximal electroshock was determined at the time of peak effect. The time of peak anticonvulsant activity was determined by administering dosages of various sizes to a group of animals and administering a maximal electroshock to the animals at intervals thereafter by supplying 60 ma. current through a corneal electrode for 0.2 second. Protection was indicated if the animal failed to show the tonic extensor component of the maximal electroshock seizure pattern in unprotected animals. The time of peak effect thus determined was used for all subsequent tests of anticonvulsant activity on the same active agent.

Anticonvulsant effectiveness was determined against a convulsive dose of Metrazol (106.25 mg/kg) injected subcutaneously; the criterior for effectiveness was failure to show clonic convulsive seizures. Anticonvulsant activity was determined against a lethal dose of strychnine sulfate (1.5 mg/kg) injected subcutaneously; the criterion was failure to die, even though seizures appeared.

Hypnotic activity or depression of the central nervous system as indicated by loss of the righting reflex (onset of sleep) was not exhibited by any dosage of any of the compounds of the present invention less than a lethal dose. Phenobarbital itself, on the other hand, did exhibit hypnotic activity using the foregoing criterion.

The following examples will serve to illustrate the practice of this invention.

EXAMPLE 1 In a 1000 ml flask equipped with a stirrer and condenser, 136.4 g. (0.536 mole) of phenobarbital sodium powder was suspended in 500 ml of dimethylformamide. The flask was placed in an ice bath. To the cold suspension was added 100 g. (1.25 mole) of chloromethyl methyl ether over a period of one hour. The resulting suspension was stirred at room temperature for 20 hours, then poured into 2000 ml of ice water and the resulting suspension was stirred for 2 hours. The crude product was removed by filtration, and washed three times with 100 ml of distilled water, then dissolved in about 500 m1 of hot ethanol. To the boiling solution 1 g. of activated carbon was added and the boiling solution was filtered through a Buchner funnel which contained a 1 cm. pressed layer of finely-divided diatomaceous silica. The cake was washed three times with 25 ml of ethanol and the hot solution together with the washings was allowed to cool to room temperature. The resulting crystals were filtered, washed with etha-' nol, and dried in a vacuum desiccator. There was obtained 58 g. of N,N'-dimethoxymethyl phenobarbital (also known as 1,3-dimethoxymethyl-5-ethyl-5-phenyl barbituric acid), melting point l15-117C.

Pharmocological testing of this compound gave the following results:

Dosage, mg/kg Acute Toxicity LD (oral) LD (intraperitoneal) Neurological Deficit TD 47 (33-66) Anticonvulsant Activity Maximal electroshock, ED 13.5 (8-23) Metrazol, ED 47 (29-75) Strychnine, ED 200 (125-320) Time of peak activity 2 hours 4 0.5 hour, but protection began to decrease after 4 hours and was nearly gone by 8 hours.

Anticonvulsant activity of the compound against maximal electroshock was also measured in the rat (Sprague Dawley Strain); the ED was found to be 6.6 (4.2-10.4).

EXAMPLE 2 5,5-Diphenylhydantoin sodium (27.5 g., 0.1 mole) was suspended in 250 ml of dimethylformamide. To the suspension was added over a period of 30 minutes 8.8 g. (8.25 ml) of chloromethyl methyl ether. The resulting suspension was stirred at room temperature overnight, then poured into 1 liter of ice water. A solid material precipitated. The suspension was stirred for one hour and the solid was removed by filtration, washed on the filter three times with ml of water, and the wet cake was dissolved in ml of ethanol with heating. To theboiling solution 1 g. of activated carbon was added and the boiling solution was filtered through a Buchner funnel containing a 1 cm pressed layer of finely divided diatomaceous silica. The cake was washed three times with 15 m1 of ethanol and the hot ethanol solution together with the washings was diluted with 100 ml of hot water, then allowed to cool to room temperature. The crystals which appeared were filtered and washed three times on the filter with 25 ml of 50% aqueous ethanol, then dried in a vacuum desiccator. There was obtained 20 g. of 3-methoxymethyl-5,5- diphenylhydantoin, m.p: 127128 (yield 67%). Pharmacological testing of this compound showed it to have LD of approximately 500 mg/kg. 1t exhibited anticonvulsant activity (maximal electroshock) within less than 0.5 hour after ingestion, and the activity lasted for more than 2 hours; the ED was less than 25 mg/kg.

EXAMPLE 3 Powdered phenobarbital sodium (25 g.) was suspended in 250 ml of dimethylformamide. With vigorous stirring 15.6 g of chloromethyl benzyl-ether was added to the suspension at room temperature. The reaction mixture was stirred overnight at room temperature. The suspension was then poured into 500 g of ice cold water, the mixture was stirred for 1 hour and the solid removed by filtration. The crude product was dissolved in about ml boiling ethanol; to the boiling solution 0.5 gram of activated carbon was added, and the boiling solution was filtered through a Buchner funnel which contained a compacted layer of finely divided diatomaceous silica. The cake was washed three times with 10 ml of ethanol and the hot solution together with the washings was allowed to cool to room temperature. The resulting crystals were filtered and recrystallized from 100 ml of ethanol in the same manner as described for the first crystallization. There was obtained 5.27 g of N,N'-dibenzyloxymethyl phenobarbital (also known as 1,3-dibenzyloxymethyl-5-ethyl- Sphenyl barbituric acid), m.p. 74-75.

EXAMPLE 4 I dissolved in hot ethanol (100 ml), 1 ,g of activated carbon was added and the boiling solution was filtered through a liluchner funnel which contained a compacted layer of finelydivided diatomaceous silica. The cake was washed three times with ml of ethanol and the hot solution together with the washings was allowe to cool to temperature. The resulting crystals were filtered and recrystallized two more times from 109 ml of ethanol in the same manner as described for the first crystallization. There was obtained 112.3 g of MhV-di-(u)butoxymethyl phenobarbital (also known as l B-di-(ni-butoxymethyl-S-ethyl-5-phenyl bnrbituric acid], mp. 7l-72C When tested as described above the compound exhibited anticorivulsant activity against maximal electroshock, the being than E00 trig/kg; and an LU (oral) greater than 500 trig/leg.

EXAMFLE 5 Phenobarbital sodium g) was dissolved in 250 ml of climethyli'ormamide at 120C. The solution was cooled at and at this temperature 25 g otchlorome thylethylether was adder. with wood stirring. The reac tion mixture was allowed to cool to room temperature and stirred for 2% hours. then poured into 560 g of ice water, stirred for 1 hour, and the solid product was removed by filtration. The solid material was dissolved 150 ml of boiling ethanol, 1 g oi activated carbon was added and the boiling solution was filtered through a Huchner funnel which contained a compacted layer of finely divided diatomaceous silica. The cake was washed three times with 10 ml of ethanol and to the hot solution together with the washings there was added 50 ml of hot water and the solution was allowed to cool to room temperature. The resulting crystals were filtered and recrystallized two more times in the same manner as described for the first crystallization. There was obtained 6.5 g of hlil'-diethoxyrnethy1 phenobarbital (also ltnown 1,3-dethoxymethyl-5-ethyl--5-pbenyl barbitu c acid}. mp. "---65. When tested as described above, the compound exhibited anticonvulsant activity against maximal electroshock, the time of peak activity i ll. hours and the being less than trig/kg.

EXAh lPlaiE s uct was removed by filtration and dissolved in ml .7

of" hot eth nol; 2 g oi activated carbon was added and the bol 3 solution was filtered through a liuehner fun 2 contained a compacted layer of finely divided titatomaceous silica. 'lhc calre was washed three times w it if} ml of ethanol and the hot solution to- Milli the washings was allowed to cool to room tern ierature. The resulting crystals were littered and rccr stalliaed two more times in the same manner as described for 'Lll? first crystallization. There was obtained oi 54o.i-butoxymethyl-fi,fi-diplii:nyllrydariloin. mp. lli8-1 20". Upon testing as described above, the compound exl'iibited anticonvulsant activity against n'laxin'ial electroshock, the ED being less than '25 o rug/kg. 'lhe Lin (oral) was also determined to be greater than 500 mg/kg.

EXAMPLE 7 5,5-Diphenylhydantoin sodium (27.5 g) was suspended 250 ml of dimethylforrnamide. To the susension was added 15 ml of chloromethylbenzylether with good stirring and the mixture was stirred at room temperature overnight. The reaction mixture was then poured into 1 liter of ice cold water and stirred for one hour. The solid product was removed by filtration and dissolved in 156 m1 of hot ethanol. To the boiling solution 1 g of activated carbon was added and the hot solution was filter d through a Buchner funnel which contained compacted layer of finely divided diatomaceous silica. The cake was washed three times with 10 mi of ethanol and the not solution together with the washings was allowed to cool to room temperature. The resulting crystals were filtered and recrystallized two more times in the same manner as described for the first crystallization. There was obtained 13.1 g of 3 henzyloxyrnethyifi.5-diphenylhydantoin, mp. 15 l--l52.

EXAMPLE 8 Powdered phenobarbital sodium (25.4 g) was suspended at room temperature in 250 ml of dimethylformamide and 22.1 g of chioromethyldodecanyl ether was added to the suspension. The reaction mixture was stirred overnight and poured into 1 liter of ice-water; the mixture was then stirred for 3 hours and the waxy solid removed by filtration. The wet solid was dissolved in 150 ml of boiling ethanol; to the hot solution 1 g of activated carbon was added and the boiling solution was filtered through a Buchner funnel which contained a compacted layer of finely divided diatomaceous silica. 'l'he filter calte was washed three times with 15 ml of ethanol and the hot solution together with the washings was allowed to cool overnight. The resulting crystals were filtered and recrystallized in the same manner as described for the first crystallization. There was obtained 13 g of i l,ll-dilauryloxymcthyl phenobarbital (also known as 1,3-dilanryloxymethyl-5-ethyl5-phenyl barbituric acid), mp. t8-50. When tested as describedahove the compound exhibited anticonvulsant activity against maximal electroshock, the peak activity occurring at 2 hours and the El) being about 50 rug/lag. The LD (oral) was determined to be greater than 5E0 mgllrg.

EXAMPLE 9 Sodium barbital (4 l g.; 0.200 mole) was suspended in 200 cc oi dimethylforrnamide in a 500 m1 flask equipped with a stirrer and condenser. Chloromethyl methyl ether (312.0 g.; 0.400 mole) was added gradually over a period of 1.5 minutes, with stirring, to the sodium best i slurry. The resulting mixture was stirred for four hours and then poured into i000 ml of ice water. Attt stirring for an additional hour, the reaction mixture was extracted with ethyl acetate. The ethyl acetate solution was dried and the ethyl acetate was evaporated under reduced pressure to provide an oil. Purification carried out by column chromatography using 604.80 mesh silica gel. lilution with a solvent mixture containing 1 part of ethyl acetate and 9 parts of benzene (by volume) yielded l,3-climethoxymethyl-5,5- tliothylbarbituric acid.

Acute Toxicity LD (oral) LD (intraperitoneal) Anticonvulsant Activity 1000 mg/kg ca 1000 mg/kg Metrazol, ED 7(4.68.4) mg/kg Time of Peak Activity 1 hour EXAMPLE 10 Sodium barbital (4.0 g.) was suspended in 50 ml of dimethylformamide. To the suspension was added, over a period of minutes, chloromethyl n-dodecyl ether g.). The suspension was heated at 1 10C. for a period of 1 hour and then allowed to stand overnight without heating. The suspension was poured into 500 ml of ice water and the resulting aqueous emulsion was extracted with ethyl acetate. The ethyl acetate solution was dried and the ethyl acetate was evaporated under reduced pressure to yield an oil, which was purified by column chromatography using silica gel (70-325 mesh). Elution with benzene provided 1,3-din-dodecyloxymethyl-S,S-diethyl barbituric acid.

Analysis calculated for C H N O C, 70.30; H, 11.11; N, 4.82; Found: C, 70.56; H, 11.17; N, 4.67

When tested as described above, this compound exhibited anticonvulsant activity against Metrazol, the ED being about 200 mg/kg; a time of peak activity of about one hour and an LD (oral) of more than 1000 mg/kg.

EXAMPLE 1 1 Sodium secobarbital (25.5 g) was suspended in dimethylformamide (250 ml'). The suspension was stirred 40 vigorously and chloromethyl methyl ether (8.5 ml) was added over a period of 10 minutes. The reaction mixture was stirred at room temperature overnight, then poured into ice water and the oily product extracted into methylene chloride. The methylene chloride solution was dried, and the methylene chloride evaporated under reduced pressure to yield a crude oil. Purification by column chromatography using silica gel (60-200 mesh) and elution with a solvent mixture containing 1 part of ethyl acetate and 9 parts of benzene (by volume) provided l,3-dimethoxymethyl-5-allyl-5- (2-pentyl) barbituric acid.

Analysis calculated for C H O N C, 58.88; H, 8.03; N, 8.58; Found: C, 58.93; H, 8.13; N, 8.60

Upon testing as previously described, the compound exhibited anticonvulsant activity against Metrazol, the ED being about 100 mg/kg and an LD (oral) of 500 mg/kg.

0 diphenylhydantoin EXAMPLE 12 Sodium secobarbital (25.5 g.) was dissolved in 250 ml of hot dimethylformamide. The solution was vigorously stirred and allowed to cool to room temperature to provide a fine suspension. To the suspension was added chloromethyl benzyl ether (16.5 g.) over a period of 10 minutes. The reaction mixture was stirred overnight and poured into 500 ml of ice water; the mixture was stirred for 1 hour and was extracted into methylene chloride. The methylene chloride solution was dried and the methylene chloride evaporated under reduced pressure to yield an oil. This crude oil was adsorbed on 60-200 mesh silica gel; elution with a solvent mixture containing 1 part of ethyl acetate and 20 parts of benzene (by volume) provided pure 1,3-dibenzyloxymethyl-5-allyl-5-(2-pentyl) barbituric acid.

Analysis calculated for C I-1 0 19 C, 70.27; H, 7.16; N, 5.85; Found: C, 70.34; H, 7.16; N, 5.67

When tested as described above, this compound exhibited anticonvulsant activity against Metrazol, the ED being about 200 mg/kg and had an LD (oral) of more than 1000 mg/kg.

What is claimed is:

1. A therapeutic composition for treatment of convulsions in warm-blooded animals comprising a physiologically acceptable carrier and an amount sufficient to produce an anticonvulsant effect of a compound selected from the group consisting of 3-alkoxymethy1- 5 ,S-diphenylhydantoin, 1,3-dialkoxymethyl-5 ,5- and 3-benzy1oxymethyl-5,5- diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.

2. The composition of claim 1 in which said compound is 3-alkoxymethy1-5,S-diphenylhydantoin in which said alkoxy contains 1 to 12 carbon atoms.

3. The composition of claim 1 in which said compound is l,3-dialkoxymethyl-5,S-diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.

4. The composition of claim 1 in which said compound is 3-benzyloxymethyl-5,S-diphenylhydantoin.

5. The composition of claim 1 in which said compound is 3-methoxymethyl-5,S-diphenylhydantoin.

6. A method of treating convulsions in warm-blooded animals which comprises administering said animals an amount sufficient to produce an anticonvulsant effect of a compound selected from the group consisting of 3-alkoxymethyl diphenylhydantoin, 1,3-dia1koxymethyl-5,5-diphenylhydantoin and 3-benzyloxymethyl-5,5- diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.

7. The method of claim 6 in which said compound is 3-a1koxymethyl-5,S-diphenylhydantoin in which said alkoxy contains 1 to 12 carbon atoms.

8. The method of claim 6 in which said compound is 1,3-dialkoxymethy1-5,5-diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.

9. The method of claim 6 in which said compound is 3-benzy1oxymethyl-5,S-diphenylhydantoin.

10. The method of claim 6 in which said compound is 3 -methoxymethyl-5 ,5 -diphenylhydantoin. 

1. A THERAPEUTIC COMPOSITION FOR TREATMENT OF CONVULSIONS IN WARM-BLOODED ANIMALS COMPRISING A PHYSIOLOGICALLY ACCEPTABLE CARRIER AND AN AMOUNT SUFFICIENT TO PRODUCE AN ANTICONVULSANT EFFECT OF A COMPOUND SELECTED FROM THE GROUP CONSISTING OF 3-ALKOXYMETHYL-5,5-DIPHENYLHYDANTOIN, 1,3-DIALKOXYMETHYL-5,5-DIPHENYLHYDANTOIN AND 3-BENZYLOXYMETHYL-5,5DIPHENYLHYDANTOIN IN WHICH EACH ALKOXY CONTAINS 1 TO 12 CARBON ATOMS.
 2. The composition of claim 1 in which said compound is 3-alkoxymethyl-5,5-diphenylhydantoin in which said alkoxy contains 1 to 12 carbon atoms.
 3. The composition of claim 1 in which said compound is 1,3-dialkoxymethyl-5,5-diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.
 4. The composition of claim 1 in which said compound is 3-benzyloxymethyl-5,5-diphenylhydantoin.
 5. The composition of claim 1 in which said compound is 3-methoxymethyl-5,5-diphenylhydantoin.
 6. A method of treating convulsions in warm-blooded animal which comprises administering said animals an amount sufficient to produce an anticonvulsant effect of a compound selected from the group consisting of 3-alkoxymethyl diphenylhydantoin, 1,3-dialkoxymethyl-5,5-diphenylhydantoin and 3-benzyloxymethyl-5,5-diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.
 7. The method of claim 6 in which said compound is 3-alkoxymethyl-5,5-diphenylhydantoin in which said alkoxy contains 1 to 12 carbon atoms.
 8. The method of claim 6 in which said compound is 1,3-dialkoxymethyl-5,5-diphenylhydantoin in which each alkoxy contains 1 to 12 carbon atoms.
 9. The method of claim 6 in which said compound is 3-benzyloxymethyl-5,5-diphenylhydantoin.
 10. The method of claim 6 in which said compound is 3-methoxymethyl-5,5-diphenylhydantoin. 